Basic underlying mechanisms of musculoskeletal aging. The long- term goal of the research is to determine the molecular mechanisms underlying diminished force generation is skeletal muscle with age. The focus of the present proposal is myosin, the major contractile protein in muscle. The hypothesis is that the decline in muscle force-generating capacity with age is due to specific structural alterations in myosin. This hypothesis will be tested by performing physiological and biophysical investigations of skeletal muscle fibers isolated from rats of different ages. The following aims will be pursued. AIM 1. Determine age-related differences in single fiber force- generating capabilities. The force-generating capacity of skeletal muscle fibers from hindlimb muscles of rats aged 3, 12, 24, 30, and 36 months will be determined. A permeabilized, single muscle fiber preparation will be used to precisely measure contractile properties of individual fibers and a sensitive microgel technique will be used to identify myosin heavy chain isoform content (i.e., fiber type will be determined). The specific hypothesis is that there will be a decline in the force- generating capacity of type II fibers with age. AIM 2. Use site- directed electron paramagnetic resonance (EPR) spectroscopy to determine age-related changes in myosin structure. Spin labels will be attached to a selected site on the myosin head within muscle fibers and then EPR will be used to detect age-related changes in myosin structure during a muscle contraction. The advantage of EPR is its high resolution, which can detect and quantitate the distribution of structural states corresponding to the weak- and strong-binding intermediates of myosin. Age- related myosin structural changes will be correlated with muscle force-generating capabilities (determined in Aim 1). The specific hypothesis that will be tested is that age-related declines in force generation result from decreases in the strong- binding structural state of myosin. These preliminary studies will be the first in a series of studies that will lead to valuable insight into one of the molecular mechanisms of aging.